Antioxidant And Anti-Diabetic Activities Of Corchorus Olitorius Linn Leaves Extract From Qassim Region, Saudi Arabia (Proposal)

Summary

Corchorus olitorius Linn. (Tiliaceae), commonly known as “Jute”, is an yearly, much-branched herb, 90-120 cm high with smooth stems, leaves of 6-10 cm long and 3. 5-5 cm wide, with light yellow flowers and black trigonous seeds. C. olitorius considered as an essential vegetable in numerous humid areas including Egypt, Sudan, India, Bangladesh, in humid Asian countries like the Philippines and Malaysia, in addition to in humid Africa, Japan, South America, the Caribbean and Cyprus. It has been reported the leaves of corchorus olitorius have medicinal utilities like wound healing, inhibition of gastric acid production and cystitis.

The aim of the study is to perform preliminary phytochemical evaluation along with quantitative total phenolic content study of C. olitorius leaves extracted by methanol maceration method. The best extracts are subjected to perform in vitro antioxidant and anti-diabetic studies. Antioxidant activity is evaluated by DPPH method and anti-diabetic property is evaluated by α-amylase enzyme inhibitory studies. Statistical analysis will be performed using Graphpad prism (version 5. 0) software. The results will be compared with corresponding standard references.

Keywords: Corchorus olitorius, maceration, antioxidant,anti-diabetic, TPC.

Background

Corchorus olitorius Linn. (Tiliaceae), commonly known as “Jute”, is an yearly, much-branched herb, 90-120 cm high with smooth stems, leaves of 6-10 cm long and 3. 5-5 cm wide, with light yellow flowers and black trigonous seeds (Kirtikar and Basu, 1987). C. olitorius considered as an essential vegetable in numerous humid areas such as Egypt, Sudan, India, Bangladesh, in humid Asian countries like the Philippines and Malaysia, in addition to in humid Africa, Japan, South America, the Caribbean and Cyprus. In some countries mostly West African countries like Ghana, Nigeria, Cameroon and Sierra Leone, the main diets consist of rice, cassava, maize, yams, and leafy vegetables are used as complement main foods (Tulio et al. , 2002). Gentle leaves of jute C. olitorius are used as a vegetable in Taiwan during the summer. The common preparation before eating is to collect the leaves and remove the stems and veins of the plant. These leaves are pressed and washed in running water to gain smashed leaves, then they cooked together with sweet potato (Yan et al. , 2013). The plant contains high protein content in its leaves, which serve as the important source for dietary protein in numerous humid countries (Al Batran et al. , 2013). C. olitorius polyphenolic isolate has been testified to have effect against obesity (Wang et. al. , 2011). Leaves extract of C. olitorius have an important wound healing activity (Barku et al. , 2013). Also, leaves extract offer protection from arsenic-induced myocardial injury (Das et al. , 2010).

Additionally, oral administration of the aqueous extract of C. olitorius inhibit gastric acid production (Owolyele et al. , 2014). Its leaves are used in many conditions like: cystitis, dysuria, fever and gonorrhe, restore the appetite and strength by its cold infusion (Adegoke & Adebayo-Tayo, 2009(. C. olitorius extract may serve as a substitute for chemical antioxidants substances due to their antioxidant ability (Ben Yakoub et al. , 2018). 2Botanical classification of the plant: Kingdom: Plantae. Clade: Angiosperms. Clade: Eudicots. Clade: Rosids. Order: Malvales. Family: Malvaceae. Genus: Corchorus. Species: C. olitorius. Literature review: A study conducted in Tunisia by (Ben Yakoub et al. , 2018) about antioxidant and antimicrobial activities of C. olitorius leaves extract. Ferric reducing antioxidant power assay and free radical scavenging activity on (DPPH) used to test antioxidant activity. They found that C. olitorius extract may serve as a substitute for chemical antioxidants substances due to their antioxidant ability and for that reason it have the possible to be bio-preservatives. Another study conducted by (Barku et al. , 2013) in Ghana. The study is about wound healing ability of C. olitorius leaves extract. Antioxidant activities of C. olitorius assessed by DPPH free radical scavenging method and wound healing ability was studied on albino rats and excision wound model. (Oboh et al. , 2012) conducted study in Nigeria about inhibitory effect of C. olitorius leaves extact on α-amylase, α-glucosidase and angiotensin I converting enzymes. The effect determination done by use α-amylase, α-glucosidase and angiotensin I converting enzyme inhibition assay. The study result stated that, the presence of polyphenols, caffeic acid, chlorogenic acid, isorhamnetin was the responsible of inhibition activity of C. olitorius leaves extract. The Lack of studies about quantification of polyphenols and also antioxidant and anti-diabetic activities of local C. olitorius leaves extract in Qassim Area, Saudi Arabia this made us to focus on the present study.

Aim and Objectives

• To collect the local Corchorus Olitorius (L. ) leaves grown in the land of Qassim Area, Saudi Arabia and extract the leaves using methanol by maceration method.
• To perform preliminary quantitative and qualitative analysis of the phytochemical content.
• To evaluate invitro antioxidant activity of Corchorus Olitorius (L. ) leaves extract by DPPH method and compare with reference standard.
• To evaluate invitro anti-diabetic property of Corchorus Olitorius (L. ) leaves extract by α-amylase inhibitory studies and compare with reference standard.

4Materials and Methods: Sample: The leaves extract of Corchorus Olitorius (L. ) will be used. Equipment: Ultraviolet spectrophotometer, Rotary shaker, Rotary evaporator. Chemicals: DPPH (2, 2-diphenyl-1-picrylhydrazyl), ethanol, ascorbic acid, absolute alcohol, Gallic acid, Ruthenium red solution, sodium carbonate, sodium nitrite, aluminium chloride, sodium hydroxide, sodium phosphate buffer, phosphate buffer: Reagents Folin-Ciocalteu reagents, Dinitrosalicylate color reagents(DNS).

Enzymes: Alpha-amylase. Methods: Extraction method: Maceration technique will be used for extraction from C. olitorius (Barku et al. , 2013). Preliminary qualitative analysis: standard biochemical testing methods for preliminary screening of phytochemicals such as alkaloids, carbohydrate, phenols, amino acids, steroids, anthocyanins, proteins, flavonoids, saponins, mucilage, gums, glycosides and tannins will be used. Quantitative analysis for phenol: Determination of total phenolic content (TPC): Phenolic concentration in the extracts will be measured using modified spectrophotometric method. The ethanolic solution of the samples with concentration of 4, 8, 12, 16 and 20 mg/mL respectively will be used in the analysis.

The reaction solution will be prepared by adding of sample (1 mL), Folin-Ciocalteu reagent (1 mL) along with 2. 5% sodium carbonate solution (2 mL). The solution will be protected for 30 minute at 37ºC. The absorbance will be measured at 750 nm. The process will be repeated for standard Gallic acid(Murthy et al. , 2011; Chigurupati et al. , 2016 ). Anti- oxidant assay: DPPH free radical scavenging assay: A quantity of 0. 44 mg DPPH will be dissolved in 1,000 mL of 95% ethanol. The solution will be preserved in dark for at least 30 minutes. In absolute alcohol at concentration of 1000, 500, 250, 100, 50, 25, and 10 μg/mL respectively, the samples and ascorbic acid (standard) will be dissolved. 500 μL of extract will be added with 500 μL DPPH solution to form sample stock.

However, 500 μL of absolute alcohol and 500 μL DPPH solutions will be added as control. Ascorbic acid will be used as standard reference (Chigurupati et al. , 2016). The calculation of radical scavenging activity will be as the percentage inhibition by using the following formula: % Inhibition = (Control Absorbance – Sample Absorbance) X 100 / Control Absorbance. Anti-diabetic assay: In Vitro anti diabetic activity Alpha amylase enzyme inhibition assay: A volume of 500 μL of sample and 500 μL acarbose at different concentrations (100, 200, 400, 800 and 1000 μg/mL respectively) will incubate with 500 μL of 0. 5 mg/mL alpha amylase in 0. 2 mM phosphate buffer (pH6. 9) at 25ºC for 10 minutes. Then, 1 % starch solution (500 μL) in 0. 02 M sodium phosphate buffer (pH 6. 9) will be added followed by incubation for 10 minutes at 25 ºC. DNS coloring reagent (1 mL) will be added and boil for 5 minutes. The test tubes will be cooled to room temperature. The reaction mixture will be diluted after adding 10 mL distilled water. The absorbance will be taken at 540 nm. The calculation of percentage of alpha amylase enzyme inhibitory will be based on the following formula (Noreen et al. , 2017).